How the variant detectors workThe Variant detectors share a set of filters. They relate to (i) which areas and positions of the read mappings that should be inspected for variants, (ii) which reads in the data should be considered when this assessment is done, (iii) requirements to the coverage, frequency and absolute counts of variant carrying reads and (iv) the quality and neighborhood composition of the area surrounding the variant. The filters are described in detail in Filters.
The variant detectors operate in the following step-wise fashion:
- Estimate an error model (Fixed Ploidy and Low Frequency Variant Detectors only).
- Examine each single nucleotide position for the presence of a potential variant while:
- Ignoring the positions, regions and reads specified by the 'Reference masking' and 'Read filter' parts of the 'General filters' (Filters).
- Applying half the cut-offs specified by the user in the 'Count and coverage filters' part of the 'General filters' (Filters).
- Applying the cut-offs specified in the Noise filters (Filters).
- For the single position variants that survived the initial screening in 2., examine if neighboring variants are present in the same reads. If so, 'join' them into MNVs, longer insertions or deletions, or into replacements.
- Apply the full cut-offs of the 'Count and coverage filters' part of the 'General filters' to all the variants (single and multiple positions) that were obtained after 3.
The reason for first examining the read mapping for variants in single positions is that it is at the single position level that we can estimate an error-model, and hence it is at the single position level that we can distinguish true variants from likely sequencing errors. The reason for joining the single position variants, when they are present in the same reads, is that this gives us evidence that they occur together (e.g., it is unsatisfactory to call three single base neighboring deletions if we can see that they occur in the same reads). The reason for applying only half of the cut-off of the 'Count and coverage filters' part of the 'General filters' on the initial single position examination (in step 2.), and wait with the full cut-off till after the variants have been joined (in step 4), is that it will sometimes happen that coverage drops within a longer variant. This can result in single position variants within a longer variant not being called, if the full filters were initially applied. By applying only half the cut-off in the initial examination, this risk is decreased.
Having described the overall mode of the variant detection tools, we will now describe individual characteristics and specific assumptions of the three variant detection tools. The filtering and output options that are shared among the tools are described in the (Filters) and Variant Detectors - the outputs.